In the detergent industry enzymes have for more than 30 years been implemented in washing formulations. Enzymes used in such formulations comprise proteases, lipases, amylases, cellulases, as well as other enzymes, or mixtures thereof. Commercially the most important enzymes are proteases.
An increasing number of commercially used proteases are protein engineered variants or naturally occurring wild type proteases, e.g. Relase®, Alcalase®, Savinase®, Primase®, Everlase®, Esperase®, Ovozyme®, Coronase®, Polarzyme® and Kannase® (Novozymes A/S), Maxatase™, Maxacal™, Maxapem™, Properase™, Purafect™, Purafect OxP™, FN2™ FN3™, FN4™ and Purafect Prime™ (Genencor International, Inc.), BLAP X and BLAP S (Henkel). Further, a number of protease variants are described in the art. A thorough list of prior art protease variants is given in WO 99/27082.
Further, a number of protease variants is described in the art, such as in EP 130 756; EP 214 435; WO 87/04461; WO 87/05050; EP 260 105; Thomas, Russell, and Fersht (1985) Nature 318:375-376; Thomas, Russell, and Fersht (1987) J. Mol. Biol. 193:803-813; Russel and Fersht Nature 328:496-500 (1987); WO 88/08028; WO 88/08033; WO 95/27049; WO 95/30011; WO 95/30010; WO 95/29979; U.S. Pat. No. 5,543,302; EP 251 446; WO 89/06279; WO 91/00345; EP 525 610; WO 94/02618.
WO 01/60963 describes detergent composition comprising variants of the B. lentus DSM 5843 alkaline protease having substitution(s) in at least one of the positions 3, 4, 99, 188, 193, 199 and 211, where the only exemplified variant is S3T+V4I+V193M+V199I+L211D. It is disclosed that these variants shows an unexpected synergistic effect in combination with certain peroxidic oxidation agents.
WO 02/088340 describes a variant of the B. lentus DSM 5843 alkaline protease having the modifications S3T+A4I+A199I+L211G. The variant shows improved performance in automatized dishwashing compared to wild type alkaline proteases.
WO 01/75087 describes subtilisin homologues that are improved for a variety of specific properties including thermal stability, activity at low temperature and alkaline stability. WO 01/68821 describes subtilase enzymes, which are suitable for removal of egg stains from for example laundry and/or hard surfaces. WO 2004/099401 describes subtilase enzymes, which are suitable for removal of egg stains from for example laundry and/or hard surfaces.
However, even though a number of useful proteases and protease variants have been described, there is still a need for further improvement of proteases or protease variants for a number of industrial uses. In particular, the problem of removing egg stains from e.g. laundry or hard surfaces has been pronounced due to the fact that substances present in the egg white inhibit many serine proteases. Therefore, an object of the present invention is to provide improved subtilase enzymes, which are suitable for removal of egg stains from for example laundry and/or hard surfaces.